Treatment of CRTH2-mediated diseases and conditions

ABSTRACT

The present invention provides indole derivatives that antagonize prostaglandin D 2 , and that can be used to treat inflammatory diseases mediated by prostaglandin D 2 .

This application is a continuation of U.S. application Ser. No.10/972,060, filed Oct. 22, 2004, which claims the benefit of GreatBritain Patent Application No. 0324763.2, filed Oct. 23, 2003. Theentirety of each of these applications is incorporated by referenceherein.

FIELD OF THE INVENTION

The present invention relates to indole derivatives that antagonizeprostaglandin D₂, and that can be used to treat inflammatory diseasesmediated by prostaglandin D₂.

BACKGROUND OF THE INVENTION

The present invention relates to the use of certain compounds in thetreatment and prevention of allergic diseases such as asthma, allergicrhinitis and atopic dermatitis and other inflammatory diseases mediatedby prostaglandin D₂ (PGD₂) acting at the CRTH2 receptor on cellsincluding eosinophils, basophils and Th2 lymphocytes.

PGD₂ is an eicosanoid, a class of chemical mediator synthesised by cellsin response to local tissue damage, normal stimuli or hormonal stimulior via cellular activation pathways. Eicosanoids bind to specific cellsurface receptors on a wide variety of tissues throughout the body andmediate various effects in these tissues. PGD₂ is known to be producedby mast cells, macrophages and Th2 lymphocytes and has been detected inhigh concentrations in the airways of asthmatic patients challenged withantigen (Murray et al, (1986), N. Engl. J. Med. 315:800-804).Instillation of PGD₂ into airways can provoke many features of theasthmatic response including bronchoconstriction (Hardy et al, (1984) N.Engl. J. Med. 311:209-213; Sampson et al, (1997) Thorax 52:513-518) andeosinophil accumulation (Emery et al, (1989) J. Appl. Physiol.67:959-962).

The potential of exogenously applied PGD₂ to induce inflammatoryresponses has been confirmed by the use of transgenic miceoverexpressing human PGD₂ synthase which exhibit exaggeratedeosinophilic lung inflammation and Th2 cytokine production in responseto antigen (Fujitani et al, (2002) J. Immunol. 168:443-449).

The first receptor specific for PGD₂ to be discovered was the DPreceptor which is linked to elevation of the intracellular levels ofcAMP. However, PGD₂ is thought to mediate much of its proinflammatoryactivity through interaction with a G protein-coupled receptor termedCRTH2 (chemoattractant receptor-homologous molecule expressed on Th2cells) which is expressed by Th2 lymphocytes, eosinophils and basophils(Hirai et al, (2001) J. Exp. Med. 193:255-261, and EP0851030 andEP-A-1211513 and Bauer et al, EP-A-1170594). It seems clear that theeffect of PGD₂ on the activation of Th2 lymphocytes and eosinophils ismediated through CRTH2 since the selective CRTH2 agonists 13,14dihydro-15-keto-PGD₂ (DK-PGD₂) and 15R-methyl-PGD₂ can elicit thisresponse and the effects of PGD₂ are blocked by an anti-CRTH2 antibody(Hirai et al, 2001; Monneret et al, (2003) J. Pharmacol. Exp. Ther.304:349-355). In contrast, the selective DP agonist BW245C does notpromote migration of Th2 lymphocytes or eosinophils (Hirai et al, 2001;Gervais et al, (2001) J. Allergy Clin. Immunol. 108:982-988). Based onthis evidence, antagonising PGD₂ at the CRTH2 receptor is an attractiveapproach to treat the inflammatory component of Th2-dependent allergicdiseases such as asthma, allergic rhinitis and atopic dermatitis.

EP-A-1170594 suggests that the method to which it relates can be used toidentify compounds which are of use in the treatment of allergic asthma,atopic dermatitis, allergic rhinitis, autoimmune disease, reperfusioninjury and a number of inflammatory conditions, all of which aremediated by the action of PGD₂ at the CRTH2 receptor.

Compounds which bind to CRTH2 are taught in WO-A-03066046 andWO-A-03066047. These compounds are not new but were first disclosed,along with similar compounds, in GB 1356834, GB 1407658 and GB 1460348,where they were said to have anti-inflammatory, analgesic andantipyretic activity. WO-A-03066046 and WO-A-03066047 teach that thecompounds to which they relate are modulators of CRTH2 receptor activityand are therefore of use in the treatment or prevention of obstructiveairway diseases such as asthma, chronic obstructive pulmonary disease(COPD) and a number of other diseases including various conditions ofbones and joints, skin and eyes, GI tract, central and peripheralnervous system and other tissues as well as allograft rejection.

PL 65781 and JP 43-24418 also relate to indole derivatives which aresimilar in structure to indomethacin and, like indomethacin, are said tohave anti-inflammatory and antipyretic activity. Thus, although this maynot have been appreciated at the time when these documents werepublished, the compounds they describe are COX inhibitors, an activitywhich is quite different from that of the compounds of the presentinvention. Indeed, COX inhibitors are contraindicated in the treatmentof many of the diseases and conditions, for example asthma andinflammatory bowel disease for which the compounds of the presentinvention are useful, although they may sometimes be used to treatarthritic conditions.

We have now discovered that certain indole derivatives in which theindole nitrogen is substituted with a carboxylic acid moiety areantagonists of PGD₂ at the CRTH2 receptor and are useful in a method forthe treatment of diseases and conditions mediated by PGD₂ at the CRTH2receptor, the method comprising administering to a patient in need ofsuch treatment a suitable amount of one of the compounds.

SUMMARY OF THE INVENTION

In one aspect the invention provides compounds of general formula (I):

wherein

-   R¹, R², R³ and R⁴ are independently hydrogen, halo, C₁-C₆ alkyl,    —O(C₁-C₆ alkyl), - —CON(R¹¹)₂, —SOR¹¹, —SO₂R¹¹, —SO₂N(R¹¹)₂,    —N(R¹¹)₂, —NR¹¹COR¹¹, —CO₂R¹¹, —COR¹¹, —SR¹¹, —OH, —NO₂ or —CN;

each R¹¹ is independently hydrogen or C₁-C₆ alkyl;

-   R⁵ and R⁶ are each independently hydrogen, or C₁-C₆ alkyl or    together with the carbon atom to which they are attached form a    C₃-C₇ cycloalkyl group;-   R⁷ is hydrogen or C₁-C₆ alkyl;-   R⁸ is an aromatic moiety optionally substituted with one or more    substituents selected from halo, C₁-C₆ alkyl, —O(C₁-C₆)alkyl,    —CON(R¹¹)₂, —SOR¹¹, —SO₂R¹¹, —SO₂N(R¹¹)₂, —N(R¹¹)₂, —NR¹¹COR¹¹,    —CO₂R¹¹, —COR¹¹, —SR¹¹, —OH, —NO₂ or —CN;

wherein R¹¹ is as defined above;

-   R⁹ is hydrogen, or C₁-C₆ alkyl;    provided that:

R⁸ is not phenyl substituted with —COOH;

when any two of R¹, R², R³ and R⁴ are hydrogen, neither of the other twoof R¹, R², R³ and R⁴ is C₃-C₆ alkyl;

or a pharmaceutically acceptable salt, hydrate, solvate, complex orprodrug thereof;

wherein the compounds are useful for the treatment or prevention ofallergic asthma, perennial allergic rhinitis, seasonal allergicrhinitis, atopic dermatitis, contact hypersensitivity (including contactdermatitis), conjunctivitis, especially allergic conjunctivitis,eosinophilic bronchitis, food allergies, eosinophilic gastroenteritis,inflammatory bowel disease, ulcerative colitis and Crohn's disease,mastocytosis and also other PGD₂-mediated diseases, for exampleautoimmune diseases such as hyper IgE syndrome and systemic lupuserythematus, psoriasis, acne, multiple sclerosis, allograft rejection,reperfusion injury, chronic obstructive pulmonary disease, as well as,in some cases, rheumatoid arthritis, psoriatic arthritis andosteoarthritis.

A second aspect of the invention provides compounds of a general formula(II):

wherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸ and R⁹ are as defined for generalformula (I); R¹² is C₁-C₆ alkyl, aryl, (CH₂)_(m)OC(═O)C₁-C₆alkyl,(CH₂)_(m)N(R¹³)₂, CH((CH₂)_(m)O(C═O)R¹⁴)₂;

m is 1 or 2;

R¹³ is hydrogen or methyl;

R¹⁴ is C₁-C₈ alkyl;

wherein compounds of formula (II) are useful for the treatment orprevention of PGD₂-mediated diseases.

A third aspect of the invention provides methods for the treatment orprevention of PGD₂-mediated diseases, the methods comprisingadministering an effective amount of one or more compounds of generalformulas (I) or (II) to a subject in need thereof.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

Therefore, in a first aspect of the invention, there is provided the useof a compound of general formula (I):

wherein

-   R¹, R², R³ and R⁴ are independently hydrogen, halo, C₁-C₆ alkyl,    —O(C₁-C₆ alkyl), - —CON(R¹¹)₂, —SOR¹¹, —SO₂R¹¹, —SO₂N(R¹¹)₂,    —N(R¹¹)₂, —NR¹¹COR¹¹, —CO₂R¹¹, —COR¹¹, —SR¹¹, —OH, —NO₂ or —CN;

each R¹¹ is independently hydrogen or C₁-C₆ alkyl;

-   R⁵ and R⁶ are each independently hydrogen, or C₁-C₆ alkyl or    together with the carbon atom to which they are attached form a    C₃-C₇ cycloalkyl group;-   R⁷ is hydrogen or C₁-C₆ alkyl;-   R⁸ is an aromatic moiety optionally substituted with one or more    substituents selected from halo, C₁-C₆ alkyl, —O(C₁-C₆)alkyl,    —CON(R¹¹)₂, —SOR¹¹, —SO₂R¹¹, —SO₂N(R¹¹)₂, —N(R¹¹)₂, —NR¹¹COR¹¹,    —CO₂R¹¹, —COR¹¹, —SR¹¹, —OH, —NO₂ or —CN;

wherein R¹¹ is as defined above;

-   R⁹ is hydrogen, or C₁-C₆ alkyl;    provided that:

R⁸ is not phenyl substituted with —COOH;

when any two of R¹, R², R³ and R⁴ are hydrogen, neither of the other twoof R¹, R², R³ and R⁴ is C₃-C₆ alkyl;

or a pharmaceutically acceptable salt, hydrate, solvate, complex orprodrug thereof;

wherein the compounds are useful for the treatment or prevention ofallergic asthma, perennial allergic rhinitis, seasonal allergicrhinitis, atopic dermatitis, contact hypersensitivity (including contactdermatitis), conjunctivitis, especially allergic conjunctivitis,eosinophilic bronchitis, food allergies, eosinophilic gastroenteritis,inflammatory bowel disease, ulcerative colitis and Crohn's disease,mastocytosis and also other PGD₂-mediated diseases, for exampleautoimmune diseases such as hyper IgE syndrome and systemic lupuserythematus, psoriasis, acne, multiple sclerosis, allograft rejection,reperfusion injury, chronic obstructive pulmonary disease, as well as,in some cases, rheumatoid arthritis, psoriatic arthritis andosteoarthritis.

WO-A-9950268, WO-A-0032180, WO-A-0151849 and WO-A-0164205 all relate tocompounds which are similar to the compounds of general formula (I).However, these compounds are said to be aldose reductase inhibitorsuseful in the treatment of diabetes mellitus (WO-A-9950268, WO-A-0032180and WO-A-0164205) or hypouricemic agents (WO-A-0151849). There is nosuggestion in any of these documents that the compounds would be usefulfor the treatment of diseases and conditions mediated by PGD₂ at theCRTH2 receptor. The preferred compounds described in these prior artdocuments mostly have a benzothiazole substituent in the positionequivalent to R⁸ of general formula (I).

U.S. Pat. No. 4,363,912 relates to compounds similar to those of thepresent invention which are said to be inhibitors of thromboxanesynthetase and to be useful in the treatment of conditions such asthrombosis, ischaemic heart disease and stroke. The compounds have apyridyl group in the position equivalent to R⁸ of general formula (I).

WO-A-9603376 relates to compounds which are said to be sPLA₂ inhibitorswhich are useful in the treatment of bronchial asthma and allergicrhinitis. These compounds all have amide or hydrazide substituents inplace of the carboxylic acid derivative of the compounds of the presentinvention.

JP 2001247570 relates to a method of producing a 3-benzothiazolylmethylindole acetic acid, which is said to be an aldose reductase inhibitor.

U.S. Pat. No. 4,859,692 relates to compounds which are said to beleukotriene antagonists useful in the treatment of conditions such asasthma, hay fever and allergic rhinitis as well as certain inflammatoryconditions such as bronchitis, atopic and ectopic eczema. The compoundsof this document are similar to the compounds of general formula (I),but general formula (I) specifically excludes compounds in which R⁸ isphenyl substituted with a —COOH group, which is the only area ofoverlap. Furthermore, J. Med. Chem., 6(33):1781-1790 (1990), which hasthe same authors as this prior patent application, teaches thatcompounds with an acetic acid group on the indole nitrogen do not havesignificant peptidoleukotriene activity. In view of this, it is mostsurprising that the compounds of the present invention, which all havean acetic acid group on the indole nitrogen, are useful for treatingconditions such as asthma, hay fever and allergic rhinitis.

U.S. Pat. No. 4,273,782 is directed to compounds similar to those ofgeneral formula (I), which are said to be useful in the treatment ofconditions such as thrombosis, ischaemic heart disease, stroke,transient ischaemic attack, migraine and the vascular complications ofdiabetes. There is no mention in the document of conditions mediated bythe action of PGD₂ at the CRTH2 receptor. The compounds of this priorart document all have an imidazole group in the position equivalent toR⁸ of general formula (I).

U.S. Pat. No. 3,557,142 relates to 3-substituted-1-indole carboxylicacids and esters which are said to be useful in the treatment ofinflammatory conditions.

WO-A-03/097598 relates to compounds which are CRTH2 receptorantagonists. They do not have an aromatic substituent in the positionequivalent to R⁸ of general formula (I).

Cross et al, J. Med. Chem. 29:342-346 (1986) relates to a process forpreparing compounds similar to those of general formula (I) from thecorresponding esters similar to the compounds of general formula (II).The compounds to which it relates are said to be inhibitors ofthromboxane synthetase and all have an imidazole group in the positionequivalent to R⁸ of general formula (I).

EP-A-0539117 relates to leukotriene antagonists which are similar instructure to the compounds of general formula (I).

US 2003/0153751 relates to compounds which are sPLA₂ inhibitors.Although the structural formula covers compounds similar to those ofgeneral formula (I), all of the exemplified compounds have bulkysubstituents at the 2- and 5-positions of the indole system and aretherefore very different from the compounds of the present invention.

US 2004/011648 discloses compounds which are similar to the compounds ofgeneral formula (I) and which are inhibitors of PAI-1. There is nosuggestion that the compounds might have CRTH2 antagonist activity.

WO 2004/058164 relates to compounds which are said to be asthma andallergic inflammation modulators. The only compounds for which activityis demonstrated are entirely different in structure from the compoundsof general formula (I).

Compounds which bind to the CRTH2 receptor are disclosed inWO-A-03/097042 and WO-A-03/097598. These compounds are indole aceticacids but in WO-A-03/097042 the indole system is fused at the 2-3positions to a 5-7 membered carbocyclic ring. In WO-A-03/097598 there isa pyrrolidine group at the indole 3-position.

WO-A-03/101981 and WO-A-03/101961 both relate to compound which are saidto be CRTH2 antagonists but which differ in structure from the compoundsof general formula (I) because there is an —S— or —SO₂— group linked tothe indole 3-position in place of the CH₂ group of the compounds ofgeneral formula (I).

In the present specification “C₁-C₆ alkyl” refers to a straight orbranched saturated hydrocarbon chain having one to six carbon atoms andoptionally substituted with one or more halo substituents or with one ormore C₃-C₇ cycloalkyl groups. Examples include methyl, ethyl, n-propyl,isopropyl, t-butyl, n-hexyl, trifluoromethyl, 2-chloroethyl,methylenecyclopropyl, methylenecyclobutyl and methylenecyclopentyl.

“C₁-C₄ alkyl” and “C₁-C₁₈ alkyl” have similar meanings except that theycontain from one to four and from one to eighteen carbon atomsrespectively.

C₃-C₇ cycloalkyl refers to a saturated 3 to 7 membered carbocyclic ring.Examples of such groups include cyclopropyl, cyclobutyl, cyclopentyl andcyclohexyl.

In the present specification, “halo” refers to fluoro, chloro, bromo oriodo.

The terms “aromatic moiety” and “aryl” in the context of the presentspecification refer to an aromatic ring system having from 5 to 14 ringcarbon atoms and containing up to three rings, one or more of which maybe replaced by a nitrogen, oxygen or sulfur atom. Examples of aromaticmoieties are benzene, pyridine, naphthalene, biphenyl, quinoline,isoquinoline, quinazoline, thiazole, benzthiazole, benzoxazole,benzimidazole, indole, indazole and imidazole ring systems.

Appropriate pharmaceutically and veterinarily acceptable salts of thecompounds of general formulae (I) and (II) include basic addition saltssuch as sodium, potassium, calcium, aluminium, zinc, magnesium and othermetal salts as well as choline, diethanolamine, ethanolamine, ethyldiamine and other well known basic addition salts.

Where appropriate, pharmaceutically or veterinarily acceptable salts mayalso include salts of organic acids, especially carboxylic acids,including but not limited to acetate, trifluoroacetate, lactate,gluconate, citrate, tartrate, maleate, malate, pantothenate, adipate,alginate, aspartate, benzoate, butyrate, digluconate, cyclopentanate,glucoheptanate, glycerophosphate, oxalate, heptanoate, hexanoate,fumarate, nicotinate, pamoate, pectinate, 3-phenylpropionate, picrate,pivalate, proprionate, tartrate, lactobionate, pivolate, camphorate,undecanoate and succinate, organic sulfonic acids such asmethanesulfonate, ethanesulfonate, 2-hydroxyethane sulfonate,camphorsulfonate, 2-naphthalenesulfonate, benzenesulfonate,p-chlorobenzenesulfonate and p-toluenesulfonate; and inorganic acidssuch as hydrochloride, hydrobromide, hydroiodide, sulfate, bisulfate,hemisulfate, thiocyanate, persulfate, phosphoric and sulfonic acids.

Salts which are not pharmaceutically or veterinarily acceptable maystill be valuable as intermediates.

Prodrugs are any covalently bonded compounds which release the activeparent drug according to general formula (I) in vivo. Examples ofprodrugs include alkyl esters of the compounds of general formula (I),for example the esters of general formula (II) below.

If a chiral centre or another form of isomeric centre is present in acompound of the present invention, all forms of such isomer or isomers,including enantiomers and diastereoisomers, are intended to be coveredherein. Compounds of the invention containing a chiral centre may beused as a racemic mixture, an enantiomerically enriched mixture, or theracemic mixture may be separated using well-known techniques and anindividual enantiomer may be used alone.

In the compounds of general formula (I), it is preferred that,independently or in any combination:

R¹ is halo or hydrogen;

R² is halo or hydrogen;

R³ is halo or hydrogen;

R⁴ is halo or hydrogen.

In more preferred compounds, R¹, R³ and R⁴ are hydrogen, while R² ishalo, particularly fluoro.

In preferred compounds of general formula (I), R⁵ and R⁶ are eachindependently hydrogen or C₁-C₄ alkyl. However, in more activecompounds, at least one, and preferably both of R⁵ and R⁶ are hydrogen.

Similarly, it is preferred that R⁹ is hydrogen or C₁-C₄ alkyl, mostpreferably hydrogen.

Compounds of general formula (I) preferably have an R⁷ group chosen fromH or C₁-C₆ alkyl; most suitably R⁷ is methyl.

In preferred compounds of general formula (I), R⁸ is phenyl,naphthalenyl, quinolinyl, quinoxalinyl, thiazolyl, biphenyl orbenzothiazolyl, any of which may optionally be substituted with one ormore substituents as defined above.

In particular, it is preferred that R⁸ is phenyl substituted at the4-position or naphthalen-2-yl, quinolin-2-yl, quinoxalin-2-yl,thiazol-2-yl or benzothiazol-2-yl, any of which may optionally besubstituted with one or more of the substituents defined above.

When the R⁸ moiety is substituted, preferred substitutents include halo,C₁-C₄ alkyl, C₁-C₄ haloalkyl, C₁-C₄ alkoxy, C₁-C₄ alkylsulfonyl andhydroxy.

Especially preferred substitutents for the R⁸ moiety include chloro,fluoro, methyl, ethyl, t-butyl, trifluoromethyl, methoxy,methanesulfonyl and hydroxy.

Among the most preferred compounds are the following:

-   1.    {3-[1-(4-Chloro-phenyl)-ethyl]-5-fluoro-2-methyl-indol-1-yl}-acetic    acid;-   2.    {5-Fluoro-2-methyl-3-[1-(4-trifluoromethyl-phenyl)-ethyl]-indol-1-yl}-acetic    acid;-   3.    {3-[1-(4-tert-Butyl-phenyl)-ethyl]-5-fluoro-2-methyl-indol-1-yl}-acetic    acid;-   4.    {5-Fluoro-3-[1-(4-methanesulfonyl-phenyl)-ethyl]-2-methyl-indol-1-yl}-acetic    acid;-   5. [5-Fluoro-2-methyl-3-(1-naphthalen-2-yl-ethyl)-indol-1-yl]-acetic    acid;-   6. (5-Fluoro-2-methyl-3-quinolin-2-ylmethyl-indol-1-yl)-acetic acid;-   7. (5-Fluoro-2-methyl-3-naphthalen-2-ylmethyl-indol-1-yl)-acetic    acid;-   8.    [5-Fluoro-3-(8-hydroxy-quinolin-2-ylmethyl)-2-methyl-indol-1-yl]-acetic    acid;-   9. (5-Fluoro-2-methyl-3-quinoxalin-2-ylmethyl-indol-1-yl)-acetic    acid;-   10. [5-Fluoro-3-(4-methoxy-benzyl)-2-methyl-indol-1-yl]-acetic acid;-   11. (5-Fluoro-2-methyl-3-thiazol-2-ylmethyl-indol-1-yl)-acetic acid    ethyl ester;-   12. [3-(4-Chloro-benzyl)-5-fluoro-2-methyl-indol-1-yl]-acetic acid;-   13. (3-Benzothiazol-2-ylmethyl-5-fluoro-2-methyl-indol-1-yl)-acetic    acid;-   14.    [5-Fluoro-2-methyl-3-(4-trifluoromethyl-benzyl)-indol-1-yl]-acetic    acid;-   15. [5-Fluoro-2-methyl-3-(4-tert-butyl-benzyl)-indol-1-yl]-acetic    acid;-   16. (3-Biphenyl-4-ylmethyl-5-fluoro-2-methyl-indol-1-yl)-acetic    acid;-   17.    [5-Fluoro-3-(4-methanesulfonyl-benzyl)-2-methyl-indol-1-yl]-acetic    acid;-   18.    [5-Fluoro-3-(6-fluoro-quinolin-2-ylmethyl)-2-methyl-indol-1-yl]-acetic    acid;-   19.    (±)-3-(1-Benzothiazol-2-yl-ethyl)-5-fluoro-2-methyl-indol-1-yl]-acetic    acid;-   20. [3-(4,5,7-Trifluoro-benzothiazol-2-ylmethyl)-indol-1-yl]-acetic    acid (lidorestat);-   21. (2-Methyl-3-quinolin-2-ylmethyl-indol-1-yl)-acetic acid;-   22. (5-Chloro-2-methyl-3-quinolin-2-ylmethyl-indol-1-yl)-acetic    acid;

or the C₁-C₆ alkyl, aryl, (CH₂)_(m)OC(═O)C₁-C₆alkyl, (CH₂)_(m)N(R³)₂,CH((CH₂)_(m)O(C═O)R¹⁴)₂ esters of any of the above; wherein

m is 1 or 2;

R¹³ is hydrogen or methyl;

R¹⁴ is C₁-C₁₈ alkyl.

Although some compounds of general formula (I) are known from the priorart, others represent a novel selection since they are not exemplifiedand the aromatic groups in the R⁸ position are not said to be preferred.Furthermore, these compounds have, surprisingly, been shown by thepresent inventors to have activity as antagonists of PGD₂ at the CRTH2receptor.

Therefore, in a further aspect of the invention there is provided acompound of general formula (I) wherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷ andR⁹ are as defined above and R⁸ is a phenyl, naphthalenyl, thiazole,biphenyl, quinolinyl or quinoxalinyl group, any of which may besubstituted with one or more halo, C₁-C₆ alkyl, —O(C₁-C₆)alkyl, —SO₂R¹¹or —OH groups;

provided that.

R⁸ is not unsubstituted phenyl or phenyl substituted with —COOH;

when any two of R¹, R², R³ and R⁴ are hydrogen, neither of the other twoof R¹, R², R³ and R⁴ is C₃-C₆ alkyl;

when all of R¹, R², R³ and R⁴ are hydrogen, R⁸ is not 4-chlorophenyl.

In these novel compounds, preferred R⁸ groups are is phenyl substitutedat the 4-position, naphthalen-2-yl, quinolin-2-yl, quinoxalin-2-yl orthiazol-2-yl and preferred substituents for these groups are chloro,fluoro, methyl, ethyl, t-butyl, trifluoromethyl, methoxy,methanesulfonyl and hydroxy.

Preferred R¹, R², R³, R⁴, R⁵, R⁶, R⁷ and R⁹ groups are as specifiedabove.

Among the most preferred novel compounds are Compounds 1 to 19, 21 and22 listed above and these compounds form a further aspect of theinvention. Compound 20 was disclosed in WO-A-9950268.

The compound of general formula (I) may be derived in vivo from aprodrug. The prodrug may be a compound of general formula (II):

wherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸ and R⁹ are as defined for generalformula (I); R¹² is C₁-C₆ alkyl, aryl, (CH₂)_(m)OC(═O)C₁-C₆alkyl,(CH₂)_(m)N(R¹³)₂, CH((CH₂)_(m)O(C═O)R¹⁴)₂;

m is 1 or 2;

R¹³ is hydrogen or methyl;

R¹⁴ is C₁-C₁₈ alkyl.

Therefore, in a further aspect of the invention there is provided theuse of a compound of general formula (II) as defined above in thepreparation of an agent for the treatment or prevention of diseases andconditions mediated by PGD₂ at the CRTH2 receptor.

Examples of particularly suitable R¹² groups when the compound ofgeneral formula (II) is used as a prodrug include:

methyl, ethyl, propyl, phenyl, CH₂C(═O)tBu, CH₂CH₂N(Me)₂, CH₂CH₂NH₂ orCH(CH₂O(C═O)R¹⁴)₂ wherein R¹⁴ is as defined above.

Compounds of general formula (II) wherein R⁸ is a phenyl, naphthalenyl,biphenyl, quinolyl or quinoxalyl group, any of which may be substitutedwith one or more halo, C₁-C₆ alkyl, —O(C₁-C₆)alkyl, —SO₂R¹¹ or —OHgroups;

provided that

R⁹ is not unsubstituted phenyl or phenyl substituted with —COOH;

when any two of R¹, R², R³ and R⁴ are hydrogen, neither of the other twoof R¹, R², R³ and R⁴ is C₃-C₆ alkyl;

when all of R¹, R², R³ and R⁴ are hydrogen, R⁸ is not 4-chlorophenyl;

are new.

Some of the most preferred compounds of general formula (II) are theC₁-C₆ alkyl, aryl, (CH₂)_(m)OC(═O)C₁-C₆alkyl, (CH₂)_(m)N(R¹³)₂,CH((CH₂)_(m)O(C═O)R¹⁴)₂ esters of Compounds 1 to 19 above, wherein m,R¹³ and R¹⁴ are as defined above.

When the compound of general formula (II) acts as a prodrug, it is latertransformed to the drug by the action of an esterase in the blood or ina tissue of the patient.

As is described in WO-A-9950268, compounds of general formula (I) may beprepared from compounds of general formula (II) in which R¹² is C₁-C₆alkyl by hydrolysis with an alkali such as sodium or lithium hydroxide.The reaction may take place in an aqueous solvent or an organic solventor a mixture of the two. A typical solvent used for the reaction is amixture of tetrahydrofuran and water.

Compounds of general formula (II) may be prepared from compounds ofgeneral formula (III):

wherein R¹, R², R³, R⁴, R⁵, R⁶ and R⁷ are as defined in general formula(I) and R¹² is as defined in general formula (II); by reaction with acompound of general formula (IV):R⁹C(═O)R⁸  (IV)wherein R⁹ is as defined for general formula (I);under acidic reductive alkylation conditions. Compounds of generalformulae (III) and (IV) are readily available or can be prepared bymethods well known to those skilled in the art.

Other methods of preparing compounds of general formulae (I) and (II)are set out in WO-A-9950268 and WO-A-0151489.

Compounds of general formula (I) are antagonists of PGD₂ at the CRTH2receptor and compounds of general formula (II) are prodrugs forcompounds of general formula (I). Compounds of general formulae (I) and(II) are therefore useful in a method for the treatment of diseases andconditions mediated by PGD₂ at the CRTH2 receptor, the method comprisingadministering to a patient in need of such treatment a suitable amountof a compound of general formula (I) or (II).

In a further aspect of the invention, there is provided a novel compoundof general formula (I) or (II) for use in medicine, particularly for usein the treatment or prevention of diseases and conditions mediated byPGD₂ at the CRTH2 receptor.

As mentioned above, such diseases and conditions include allergicasthma, perennial allergic rhinitis, seasonal allergic rhinitis, atopicdermatitis, contact hypersensitivity (including contact dermatitis),conjunctivitis, especially allergic conjunctivitis, eosinophilicbronchitis, food allergies, eosinophilic gastroenteritis, inflammatorybowel disease, ulcerative colitis and Crohn's disease, mastocytosis andalso other PGD₂-mediated diseases, for example autoimmune diseases suchas hyper IgE syndrome and systemic lupus erythematus, psoriasis, acne,multiple sclerosis, allograft rejection, reperfusion injury, chronicobstructive pulmonary disease, as well as rheumatoid arthritis,psoriatic arthritis and osteoarthritis.

The compounds of general formula (I) or (II) must be formulated in anappropriate manner depending upon the diseases or conditions they arerequired to treat.

Therefore, in a further aspect of the invention there is provided apharmaceutical composition comprising a novel compound of generalformula (I) or (II) together with a pharmaceutical excipient or carrier.Other active materials may also be present, as may be consideredappropriate or advisable for the disease or condition being treated orprevented.

The carrier, or, if more than one be present, each of the carriers, mustbe acceptable in the sense of being compatible with the otheringredients of the formulation and not deleterious to the recipient.

The formulations include those suitable for oral, rectal, nasal,bronchial (inhaled), topical (including eye drops, buccal andsublingual), vaginal or parenteral (including subcutaneous,intramuscular, intravenous and intradermal) administration and may beprepared by any methods well known in the art of pharmacy.

The route of administration will depend upon the condition to be treatedbut preferred compositions are formulated for oral, nasal, bronchial ortopical administration.

The composition may be prepared by bringing into association the abovedefined active agent with the carrier. In general, the formulations areprepared by uniformly and intimately bringing into association theactive agent with liquid carriers or finely divided solid carriers orboth, and then if necessary shaping the product. The invention extendsto methods for preparing a pharmaceutical composition comprisingbringing a novel compound of general formula (I) or (II) in conjunctionor association with a pharmaceutically or veterinarily acceptablecarrier or vehicle.

Formulations for oral administration in the present invention may bepresented as: discrete units such as capsules, sachets or tablets eachcontaining a predetermined amount of the active agent; as a powder orgranules; as a solution or a suspension of the active agent in anaqueous liquid or a non-aqueous liquid; or as an oil-in-water liquidemulsion or a water in oil liquid emulsion; or as a bolus etc.

For compositions for oral administration (e.g. tablets and capsules),the term “acceptable carrier” includes vehicles such as commonexcipients e.g. binding agents, for example syrup, acacia, gelatin,sorbitol, tragacanth, polyvinylpyrrolidone (Povidone), methylcellulose,ethylcellulose, sodium carboxymethylcellulose,hydroxypropylmethylcellulose, sucrose and starch; fillers and carriers,for example corn starch, gelatin, lactose, sucrose, microcrystallinecellulose, kaolin, mannitol, dicalcium phosphate, sodium chloride andalginic acid; and lubricants such as magnesium stearate, sodium stearateand other metallic stearates, glycerol stearate stearic acid, siliconefluid, talc waxes, oils and colloidal silica. Flavouring agents such aspeppermint, oil of wintergreen, cherry flavouring and the like can alsobe used. It may be desirable to add a colouring agent to make the dosageform readily identifiable. Tablets may also be coated by methods wellknown in the art.

A tablet may be made by compression or moulding, optionally with one ormore accessory ingredients. Compressed tablets may be prepared bycompressing in a suitable machine the active agent in a free flowingform such as a powder or granules, optionally mixed with a binder,lubricant, inert diluent, preservative, surface-active or dispersingagent. Moulded tablets may be made by moulding in a suitable machine amixture of the powdered compound moistened with an inert liquid diluent.The tablets may optionally be coated or scored and may be formulated soas to provide slow or controlled release of the active agent.

Other formulations suitable for oral administration include lozengescomprising the active agent in a flavoured base, usually sucrose andacacia or tragacanth; pastilles comprising the active agent in an inertbase such as gelatin and glycerin, or sucrose and acacia; andmouthwashes comprising the active agent in a suitable liquid carrier.

For topical application to the skin, compounds of general formula (I) or(II) may be made up into a cream, ointment, jelly, solution orsuspension etc. Cream or ointment formulations that may be used for thedrug are conventional formulations well known in the art, for example,as described in standard text books of pharmaceutics such as the BritishPharmacopoeia.

Compounds of general formula (I) or (II) may be used for the treatmentof the respiratory tract by nasal, bronchial or buccal administrationof, for example, aerosols or sprays which can disperse thepharmacological active ingredient in the form of a powder or in the formof drops of a solution or suspension. Pharmaceutical compositions withpowder-dispersing properties usually contain, in addition to the activeingredient, a liquid propellant with a boiling point below roomtemperature and, if desired, adjuncts, such as liquid or solid non-ionicor anionic surfactants and/or diluents. Pharmaceutical compositions inwhich the pharmacological active ingredient is in solution contain, inaddition to this, a suitable propellant, and furthermore, if necessary,an additional solvent and/or a stabiliser. Instead of the propellant,compressed air can also be used, it being possible for this to beproduced as required by means of a suitable compression and expansiondevice.

Parenteral formulations will generally be sterile.

Typically, the dose of the compound will be about 0.01 to 100 mg/kg; soas to maintain the concentration of drug in the plasma at aconcentration effective to inhibit PGD₂ at the CRTH2 receptor. Theprecise amount of a compound of general formula (I) or (II) which istherapeutically effective, and the route by which such compound is bestadministered, is readily determined by one of ordinary skill in the artby comparing the blood level of the agent to the concentration requiredto have a therapeutic effect.

Compounds of general formula (I) or (II) may be used in combination withone or more active agents which are useful in the treatment of thediseases and conditions listed above, although these active agents arenot necessarily inhibitors of PGD₂ at the CRTH2 receptor.

Therefore, the pharmaceutical composition described above mayadditionally contain one or more of these active agents.

There is also provided the use of a compound of general formula (I) or(II) in the preparation of an agent for the treatment of diseases andconditions mediated by PGD₂ at the CRTH2 receptor, wherein the agentalso comprises an additional active agent useful for the treatment ofthe same diseases and conditions.

These additional active agents which may have a completely differentmode of action include existing therapies for allergic and otherinflammatory diseases including:

β2 agonists such as salmeterol;

corticosteroids such as fluticasone;

antihistamines such as loratidine;

leukotriene antagonists such as montelukast;

anti-IgE antibody therapies such as omalizumab;

anti-infectives such as fusidic acid (particularly for the treatment ofatopic dermatitis);

anti-fungals such as clotrimazole (particularly for the treatment ofatopic dermatitis);

immunosuppressants such as tacrolimus and particularly pimecrolimus inthe case of inflammatory skin disease.

CRTH2 antagonists may also be combined with therapies that are indevelopment for inflammatory indications including:

other antagonists of PGD₂ acting at other receptors, such as DPantagonists;

inhibitors of phosphodiesterase type 4 such as cilonilast;

drugs that modulate cytokine production such as inhibitors of TNFαconverting enzyme (TACE);

drugs that modulate the activity of Th2 cytokines IL-4 and IL-5 such asblocking monoclonal antibodies and soluble receptors;

PPAR-γ agonists such as rosiglitazone;

5-lipoxygenase inhibitors such as zileuton.

In yet a further aspect of the invention, there is provided a productcomprising a novel compound of general formula (I) or (II) and one ormore of the agents listed above as a combined preparation forsimultaneous, separate or sequential use in the treatment of a diseaseor condition mediated by the action of PGD₂ at the CRTH2 receptor.

The invention will now be described in greater detail with reference tothe following non limiting examples.

EXAMPLES Example 1

Preparation of Compounds 1 to 19

1. {3-[1-(4-Chloro-phenyl)-ethyl]-5-fluoro-2-methyl-indol-1-yl}-aceticacid ethyl ester

Triethylsilane (0.34 ml, 2.13 mmol) and trifluoroacetic acid (0.10 ml,1.29 mmol) were sequentially added dropwise over 1 min to a stirredsolution of (5-fluoro-2-methyl-indol-1-yl)-acetic acid ethyl ester (0.10g, 0.43 mmol) and 4-acetylchlorobenzenze (64 mg, 0.41 mmol) in1,2-dichloroethane (2 ml) at 0° C. The mixture was then warmed to roomtemperature and stirred for 16 h. The resulting mixture was concentratedin vacuo to leave a residue which was partitioned between ethyl acetate(10 ml) and a saturated solution of sodium bicarbonate (10 ml). Theorganic layer was separated, dried, and concentrated in vacuo to leave aresidue which was purified by flash column chromatography (Flashmaster)on silica gel eluting with 10% ethyl acetate:heptane to 25% ethylacetate:heptane to give the ethyl ester (57 mg, 37%) as a white solid,Tr=1.88 min (92%), m/z (ES⁺) (M+H)⁺374.30.

2. Compound1-{3-[1-(4-Chloro-phenyl)-ethyl]-5-fluoro-2-methyl-indol-1-yl}-aceticacid

Lithium hydroxide monohydrate (70 mg, 1.67 mmol) was added in oneportion to a stirred solution of{3-[1-(4-chloro-phenyl)-ethyl]-5-fluoro-2-methyl-indol-1-yl}-acetic acidethyl ester (57 mg, 0.15 mmol) in tetrahydrofuran:water (5 ml; 1:1) andstirred at room temperature for 2 h. The solution was adjusted to pH 1with concentrated hydrochloric acid and the product extracted with ethylacetate (3×10 ml). The combined organic extracts were dried andconcentrated in vacuo to give the carboxylic acid (35 mg, 67%) as anoff-white solid, δ_(H) (400 MHz, CDCl₃) 7.26-7.21 (4H, m, Ar), 7.06 (1H,dd J 9.0, 4.2 Hz, Ar), 6.97 (1H, dd J 10.0, 2.4 Hz, Ar), 6.86 (1H, dt J9.0, 2.4 Hz, Ar), 4.80 (2H, s, CH₂CO₂H), 4.35 (1H, q J 7.3 Hz, CHCH₃),2.29 (3H, s, CH₃), 1.73 (3H, d J 7.3 Hz, CHCH₃); Tr=1.73 min (90%), m/z(ES⁺) (M+H)⁺346.09.

Compounds 2 to 19 were prepared using a similar method to that describedfor Compound 1, but with appropriately chosen starting materials.

Compound2-{5-Fluoro-2-methyl-3-[1-(4-trifluoromethyl-phenyl)-ethyl]-indol-1-yl}-aceticacid

δ_(H) (400 MHz, CDCl₃) 7.50 (2H, d J 8.3 Hz, Ar), 7.39 (2H, d J 8.3 Hz,Ar), 7.07 (1H, dd J 8.8, 4.1 Hz, Ar), 6.98 (1H, dd J 10.0, 2.5 Hz, Ar),6.85 (1H, dt J 9.0, 2.5 Hz, Ar), 4.80 (2H, s, CH₂CO₂H), 4.42 (1H, q J7.1 Hz, CHCH₃), 2.29 (3H, s, CH₃), 1.77 (3H, d J 7.3 Hz, CHCH₃); Tr=1.65min (96%), m/z (ES⁺) (M+H)⁺ 380.15.

Compound3-{3-[1-(4-tert-Butyl-phenyl)-ethyl]-5-fluoro-2-methyl-indol-1-yl}-aceticacid

δ_(H) (400 MHz, CDCl₃) 7.32-7.21 (4H, m, Ar), 7.08-7.03 (2H, m, Ar),6.89-6.83 (1H, m, Ar), 4.82 (2H, s, CH₂CO₂H), 4.36 (1H, q J 7.3 Hz,CHCH₃), 2.33 (3H, s, CH₃), 1.75 (3H, d J 7.3 Hz, CHCH₃), 1.29 (9H, s,C(CH₃)₃); Tr=1.78 min (97%), m/z (ES⁺) (M+H)⁺ 368.21.

Compound4-{5-Fluoro-3-[1-(4-methanesulfonyl-phenyl)-ethyl]-2-methyl-indol-1-yl}-aceticacid

δ_(H) (400 MHz, CDCl₃) 7.81 (2H, d J 8.3 Hz, Ar), 7.47 (2H, d J 8.1 Hz,Ar), 7.06 (1H, dd J 8.8, 4.1 Hz, Ar), 6.96 (1H, dd J 10.0, 2.5 Hz, Ar),6.85 (1H, dt J 9.0, 2.5 Hz, Ar), 4.78 (2H, s, CH₂CO₂H), 4.43 (1H, q J7.1 Hz, CHCH₃), 2.99 (3H, s, CH₃), 2.29 (3H, s, CH₃), 1.79 (3H, d J 7.3Hz, CHCH₃); Tr=1.34 min (100%), m/z (ES⁺) (M+H)⁺390.16.

Compound5-[5-Fluoro-2-methyl-3-(1-naphthalen-2-yl-ethyl)-indol-1-yl]-acetic acid

δ_(H) (400 MHz, CDCl₃) 7.81-7.74 (3H, m, Ar), 7.69 (1H, d J 8.5 Hz, Ar),7.47-7.39 (2H, m, Ar), 7.39-7.33 (1H, m, Ar), 7.09-7.02 (2H, m, Ar),6.86 (1H, dt J 9.0, 2.4 Hz, Ar), 4.83 (2H, s, CH₂CO₂H), 4.54 (1H, q J7.3 Hz, CHCH₃), 2.32 (3H, s, CH₃), 1.86 (3H, d J 7.3 Hz, CHCH₃); Tr=1.66min (97%), m/z (ES⁺) (M+H)⁺362.19.

Compound 6-(5-Fluoro-2-methyl-3-quinolin-2-ylmethyl-indol-1-yl)-aceticacid

δ_(H) (400 MHz, d₆-DMSO) 8.42 (1H, d J 9.0 Hz, Ar), 8.23 (1H, d J 9.0Hz, Ar), 8.11 (1H, m, Ar), 7.97 (1H, m, Ar), 7.60 (1H, m, Ar) 7.51 (3H,m, Ar and Ar), 7.09 (1H, m, Ar), 5.19 (2H, s, CH₂), 4.56 (2H, CH₂), 2.63(3H, s, CH₃); Tr=1.06 min (100%), m/z (ES⁺) (M+H)⁺349.35.

Compound 7-(5-Fluoro-2-methyl-3-naphthalen-2-ylmethyl-indol-1-yl)-aceticacid

δ_(H) (400 MHz, d₆-DMSO) 7.87 (4H, m, Ar), 7.47 (4H, m, Ar), 7.22 (1H,dd J 6.0, 1.5 Hz, Ar), 6.91 (1H, ddd J 9.0, 2.5 Hz, Ar), 5.04 (2H, s,CH₂), 4.23 (2H, s, CH₂), 2.42 (3H, s, CH₃); Tr=2.09 min, m/z (ES⁺)(M+H)⁺348.13.

Compound8-[5-Fluoro-3-(8-hydroxy-quinolin-2-ylmethyl)-2-methyl-indol-1-yl]-aceticacid

δ_(H) (400 MHz, d₆-DMSO) 9.53 (1H, s, OH), 8.20 (1H, d J 8.0 Hz, Ar),7.42 (5H, m, Ar), 7.13 (1H, dd J 6.0, 1.5 Hz, Ar), 6.91 (1H, dd J 9.0,2.5 Hz, Ar), 5.00 (2H, s, CH₂), 4.41 (2H, s, CH₂), 2.47 (3H, s, CH₃);Tr=1.13 min, m/z (ES⁺) (M+H)⁺365.12.

Compound 9-(5-Fluoro-2-methyl-3-quinoxalin-2-ylmethyl-indol-1-yl)-aceticacid

δ_(H) (400 MHz, d₆-DMSO) 9.02 (1H, s, H-3 Ar), 8.30 (2H, m, Ar), 8.05(2H, m, Ar), 7.53 (2H, m, Ar), 7.07 (1H, m, Ar), 5.01 (2H, br s, CH₂),4.64 (2H, s, CH₂), 2.64 (3H, s, CH₃); Tr=1.35 min, m/z (ES⁺)(M+H)⁺350.12.

Compound 10-[5-Fluoro-3-(4-methoxy-benzyl)-2-methyl-indol-1-yl]-aceticacid

δ_(H) (400 MHz, d₆-DMSO) 7.39 (1H, m, Ar), 7.16 (3H, m, Ar), 6.91 (3H,m, Ar) 5.00 (2H, s, CH₂), 3.98 (2H, s, CH₂), 3.74 (3H, s, OCH₃) 2.36(3H, s, CH₃); Tr=1.93 min, m/z (ES⁺) (M+H)⁺328.13.

Compound 11-(5-Fluoro-2-methyl-3-thiazol-2-ylmethyl-indol-1-yl)-aceticacid ethyl ester

Tr=1.09 min, m/z (ES⁺) (M+H)⁺305.26.

Compound 12-[3-(4-Chloro-benzyl)-5-fluoro-2-methyl-indol-1-yl]-aceticacid

Tr=1.63 min(100%), m/z (ES⁺) (M+H)⁺332.16.

Compound13-(3-Benzothiazol-2-ylmethyl-5-fluoro-2-methyl-indol-1-yl)-acetic acid

Tr=1.43 min, m/z (ES⁺) (M+H)⁺355.17.

Compound14-[5-Fluoro-2-methyl-3-(4-trifluoromethyl-benzyl)-indol-1-yl]-aceticacid

Tr=1.66 min, m/z (ES⁺) (M+H)⁺366.06.

Compound15-[5-Fluoro-2-methyl-3-(4-tert-butyl-benzyl)-indol-1-yl]-acetic acid

Tr=1.73 min, m/z (ES⁺) (M+H)⁺354.21.

Compound 16-(3-Biphenyl-4-ylmethyl-5-fluoro-2-methyl-indol-1-yl)-aceticacid

Tr=2.10 min, m/z (ES⁺) (M+H)⁺374.16.

Compound17-[5-Fluoro-3-(4-methanesulfonyl-benzyl)-2-methyl-indol-1-yl]-aceticacid

Tr=1.35 min, m/z (ES⁺)=376.05.

Compound18-[5-Fluoro-3-(6-fluoro-quinolin-2-ylmethyl)-2-methyl-indol-1-yl]-aceticacid

δ_(H) (400 MHz, d₆-DMSO) 8.20 (1H, d J 8.6 Hz, Ar), 8.06 (1H, dd J 9.3,5.6 Hz, Ar), 7.70 (1H, dd J 9.4, 2.8 Hz, Ar), 7.64 (1H, td J 8.8, 2.9Hz, Ar), 7.37-7.32 (2H, m, Ar), 7.26 (1H, dd J 9.9, 2.6 Hz, Ar), 6.86(1H, td J 9.2, 2.4 Hz, Ar), 4.94 (2H, s, CH₂), 4.33 (2H, s, CH₂), 2.40(3H, s, CCH₃); Tr=1.28 min (100%), m/z (ES⁺) (M+H)⁺367.50.

Compound19-(±)-3-(1-Benzothiazol-2-yl-ethyl)-5-fluoro-2-methyl-indol-1-yl]-aceticacid

δ_(H) (400 MHz, d₆-DMSO) 8.01 (1H, d J 7.7 Hz, Ar), 7.95 (1H, d J 8.0Hz, Ar), 7.49 (1H, obs t J 7.2 Hz, Ar), 7.43-7.36 (2H, m, Ar), 7.10 (1H,dd J 10.1, 2.5 Hz, Ar), 6.89 (1H, td J 9.2, 2.4 Hz, Ar), 5.01 (2H, s,CH₂), 4.91 (1H, q J 7.1 Hz, CHCH₃), 2.37 (3H, s, CCH₃), 1.87 (3H, d J7.1 Hz, CHCH₃); Tr=1.53 min, m/z (ES⁺) (M+H)⁺369.10.

Compound 21-(2-Methyl-3-quinolin-2-ylmethyl-indol-1-yl)-acetic acid

δ_(H) (400 MHz, d₆-DMSO) 8.16 (1H, d J 8.6 Hz, Ar), 8.01 (1H, d J 8.5Hz, Ar), 7.88 (1H, d J 7.6 Hz, Ar), 7.74 (1H, t J 7.0 Hz, Ar), 7.54 (1H,t J 7.0 Hz, Ar), 7.44 (1H, d J 8.0 Hz, Ar), 7.26 (2H, app t J 8.9 Hz,Ar), 7.00 (1H, t J 7.3 Hz, Ar), 6.90 (1H, t J 7.3 Hz, Ar), 4.72 (2H, s,CH₂CO₂H), 4.35 (2H, s, CH₂), 2.40 (3H, s, CH₃); Tr=1.07 min (100%), m/z(ES⁺) (M+H)⁺331.33.

Compound 22-(5-Chloro-2-methyl-3-quinolin-2-ylmethyl-indol-1-yl)-aceticacid

δ_(H) (400 MHz, d₆-DMSO) 8.21 (1H, d J 8.4 Hz, Ar), 8.00 (1H, d J 8.4Hz, Ar), 7.89 (1H, d J 8.0 Hz, Ar), 7.77-7.73 (1H, m, Ar), 7.57-7.53(2H, m, Ar), 7.40 (1H, d J 8.7 Hz, Ar), 7.29 (1H, d J 8.5 Hz, Ar), 7.04(1H, dd J 8.6, 2.1 Hz, Ar), 5.00 (2H, s, CH₂CO₂H), 4.35 (2H, s, CH₂),2.41 (3H, s, CH₃); Tr=1.17 min (95%), m/z (ES⁺) (M+H)⁺365.28.

EXAMPLE 2 Preparation of Compound 20

1.[3-(4,5,7-Trifluoro-benzothiazol-2-ylmethyl)-indol-1-yl]-acetic acidethyl ester

This compound was prepared using the procedure set out in WO-A-0151489.

δ_(H) (400 MHz, d₆-DMSO) 7.75-7.69 (1H, m, Ar), 7.56 (1H, d J 7.8 Hz,Ar), 7.49 (1H, s, CH), 7.43 (1H, d J 8.2 Hz, Ar), 7.19 (1H, app t J 7.0Hz, Ar), 7.08 (1H, app t J 7.1 Hz, Ar), 5.17 (2H, s, CH₂), 4.69 (2H, s,CH₂); 4.17 (2H, q J 7.2 Hz, CH₂CH₃), 1.23 (3H, t J 7.2 Hz, CH₂CH₃);Tr=1.62 min, m/z (ES⁺) (M+H)⁺405.15.

2. Compound 20-[3-(4,5,7-Trifluoro-benzothiazol-2-ylmethyl)-indol-1-yl]-acetic acid

Lithium hydroxide (31 mg, 0.74 mmol) in water (6 ml) was added in oneportion to a stirred solution of[3-(4,5,7-trifluoro-benzothiazol-2-ylmethyl)-indol-1-yl]-acetic acidethyl ester (73 mg, 0.18 mmol) in tetrahydrofuran (6 ml) at roomtemperature. The mixture was stirred at room temperature for 15 min andthen the solution was adjusted to pH ˜3 with 1M hydrochloric acid. Theaqueous layer was then extracted with ethyl acetate (3×10 ml) and thecombined organic extracts were washed with brine (10 ml), dried andconcentrated in vacuo to give the carboxylic acid (62 mg, 92%) as ayellow solid, δ_(H) (400 MHz, d₆-DMSO) 7.76-7.69 (1H, m, Ar), 7.56 (1H,d J 8.0 Hz, Ar), 7.48 (1H, s, CH), 7.43 (1H, d J 8.3 Hz, Ar), 7.18 (1H,app t J 7.1 Hz, Ar), 7.07 (1H, app t J 7.1 Hz, Ar), 5.05 (2H, s, CH₂),4.68 (2H, s, CH₂); Tr=1.94 min, m/z (ES⁺) (M+H)⁺377.00.

EXAMPLE 3 Measurement of CRTH2 Antagonist Activity

-   -   Materials and Methods

Materials

Calcium-3 dye was purchased from Molecular Devices (Wokingham, UK).Mono-.poly resolving medium was obtained from Dainippon Pharmaceuticals(Osaka, Japan). Macs anti-CD16 microbeads were from Miltenyi biotec(Bisley, Surrey). ChemoTx plates were purchased from Neuroprobe(Gaithersburg, Md.). Poly-D-lysine coated 96-well plates were obtainedfrom Greiner (Gloucestershire, UK). [³H]PGD₂ was from AmershamBiosciences (Buckinghamshire, UK). [³H]SQ29548 was purchased from PerkinElmer Life Sciences (Buckinghamshire, UK). All other reagents wereobtained from Sigma-Aldrich (Dorset, UK), unless otherwise stated.

Methods

-   -   Cell Culture

Chinese Hamster Ovary cells were transfected with CRTH2 or DP receptors(CHO/CRTH2 and CHO/DP) and were maintained in culture in a humidifiedatmosphere at 37° C. (5% CO₂) in Minimum Essential Medium (MEM)supplemented with 10% foetal bovine serum, 2 mM glutamine, and 1 mg ml⁻¹active G418. The cells were passaged every 2-3 days. For radioligandbinding assay, cells were prepared in triple-layer flasks or in 175 cm²square flasks (for membrane preparation). For calcium mobilisationassay, cells were grown in a 96 well plate 24 h prior to the assay at adensity of 80,000 cells per well.

-   -   Preparation of Cell Membranes

Membranes were prepared either from CHO/CRTH2 and CHO/DP cells, or fromplatelets (as a source of TP receptors). CHO cells grown to confluencywere washed with PBS and detached using a Versene solution (15 ml perflask). When the cells were grown in 175 cm² square flask, they werecollected by scrapping in PBS. The cell suspensions were centrifuged(1,700 rpm, 10 min, 4° C.) and resuspended in 15 ml of buffer (1×HBSS,supplemented with 10 mM HEPES, pH 7.3). Cell suspensions were thenhomogenised using an Ultra Turrax at setting 4-6 for 20 s. Thehomogenate was centrifuged at 1,700 rpm for 10 min and the supernatantwas collected and centrifuged at 20,000 rpm for 1 h at 4° C. Theresulting pellet was resuspended in buffer and stored at −80° C. inaliquots of 200-500 μl. The protein concentration was determined by themethod of Bradford (1976), using bovine serum albumin as standard. Theplatelets were washed by centrifugation at 600×g for 10 min andresuspended in ice-cold assay buffer (10 mM Tris-HCl, pH 7.4, 5 mMGlucose, 120 mM NaCl, 10 μM indomethacin) and directly centrifuged at20,000 rpm for 30 min at 4° C. The resulting pellet was treated asdescribed above.

-   -   Radioligand Binding Assays

[³H]PGD₂ (160 Ci/mmol) binding experiments were performed on membranesprepared as described above. Assays were performed in a final volume of100 μl of buffer (1×HBSS/HEPES 10 mM, pH 7.3). Cell membranes (15 μg).Cell membranes 15 mg were preincubated at room temperature with varyingconcentration of competing ligand for 15 min. [³H]PGD₂ (mol, finalconcentration) was then added and the incubation continued for a furtherone hour at room temperature. The reaction was terminated by theaddition of 200 μl ice-cold assay buffer to each well, followed by rapidfiltration through Whatman GF/B glass fibre filters using a UnifilterCell harvester (PerkinElmer Life Sciences) and six washes of 300 μl ofice-cold buffer. The Unifilter plates were dried at room temperature forat least 1 h and the radioactivity retained on the filters wasdetermined on a Beta Trilux counter (PerkinElmer Life Sciences),following addition of 40 μl of Optiphase Hi-Safe 3 (Wallac) liquidscintillation. Non specific binding was defined in the presence of 10 μMunlabelled PGD₂. Assays were performed in duplicate.

The results of the radioligand binding experiments to the CRTH2 and DPreceptors are shown in Tables 1 and 2.

TABLE 1 Radioligand binding data (Ki on CRTH2 Receptor). Compounds Ki(nM) Compound 4 5 ± 4 Compound 6 9 ± 3 Compound 8 6 ± 4 Compound 12 11 ±2  Compound 13 6 ± 1 Compound 17 7 ± 2 Compound 18 1.3 ± 0.6 Compound 20(lidorestat) 886 ± 248

TABLE 2 Radioligand binding data (Ki on DP Receptor). Compounds Ki (nM)Compound 4 30440 ± 9805  Compound 6 17870 ± 7290  Compound 8 7710 ± 1780Compound 12 12220 ± 2250  Compound 18 7740 ± 1442 Compound 20(lidorestat) 3960

The TP receptor radioligand binding was done on membranes prepared fromplatelets. 15-40 μg of protein were pre-incubated with varyingconcentrations of competing ligand for 15 min at room temperature inassay buffer (10 mM Tris-HCl, pH 7.4, 5 mM glucose, 120 mM NaCl, 10 μMindomethacin). [³H]SQ29548 (38 Ci/mmol, 10 nM final concentration) wasthen added and the incubation continued for a further 30 min at roomtemperature. The reaction was terminated by the addition of 200 μlice-cold assay buffer to each well, followed by rapid filtration throughWhatman GF/C glass fibre filters using a Unifilter Cell harvester(PerkinElmer Life Sciences) followed with six washes of 300 μl ofice-cold buffer. The radioactivity was determined as described above.

All of the compounds studied in this assay bound to the TP receptor withlow affinity (Ki>1 μM).

Compounds of general formula (I) bound to CRTH2 receptor expressed inCHO cells with a range of affinity varying from very high to moderate.In fact the Ki values determined in competition versus [³H]PGD₂ variedfrom 500 μM to 1 μM. Compounds of general formula (I) had no activity(or very weak activity) at the DP and TP receptors. The bindingselectivity of the compounds of general formula (I) for CRTH2 receptorwas greater than 200 fold for CRTH2 receptor, compared to DP and TPreceptors.

-   -   Calcium Mobilisation Assay

Cells were seeded onto poly-D-lysine coated 96-well plates at a densityof 80,000 cells per well and incubated at 37° C. overnight to allow thecells to adhere. Cells were washed twice with HBSS and incubated for 1 hat 37° C. in 100 μl HBSS and 100 μl calcium-3-dye (Molecular Devices),supplemented with 4 mM probenecid. Changes in fluorescence weremonitored over a 50 s time course with agonist addition at 17 s using aFlexstation (Molecular Devices).

-   -   Effect of CRTH2 Agonists on Calcium Mobilisation in CHO—CRTH2        Cells

PGD₂ caused a dose-dependent increase in intracellular Ca²⁺ mobilisationin CHO/CRTH2 cells, with an EC₅₀=2.4±0.5 nM (n=3).

-   -   Effect of Compounds of General Formula (I) on the Calcium        Mobilisation Induced by PGD₂

PGD₂-stimulated Ca²⁺ flux was fully inhibited by the compounds ofgeneral formula (I) and the IC₅₀ value for each compound in the calciumassay was comparable to its Ki value in Radioligand binding. IC₅₀ valuesof compounds of general formula (I) varied from 5 nM to 1 μM. Theresults for several compounds of general formula (I) are shown in Table3. Increasing doses of the compounds of general formula (I) caused adose-dependent and parallel shift of the PGD₂ dose response curve inCHO/CRTH2 cells, thereby indicating that the compounds are competitiveCRTH2 antagonists.

The antagonistic effect of the compounds of general formula (I) appearsto be CRTH2 selective, since no inhibitory effect was seen withATP-stimulated Ca²⁺ flux.

TABLE 3 Inhibition of PGD₂-induced calcium flux Compounds IC₅₀ (nM)Compound 4 55 ± 18 Compound 6 30 ± 6  Compound 7 38 ± 16 Compound 8 11 ±6  Compound 10 47 ± 8  Compound 12 108 ± 29  Compound 17 64 ± 5 Compound 18 10 ± 5  Compound 19 34 ± 7  Compound 20 (lidorestat) 885 ±96 

While the preferred embodiment of the invention has been illustrated anddescribed, it will be appreciated that various changes can be madetherein without departing from the spirit and scope of the invention.

1. A process for preparation of a compound of general formula (I)

wherein R¹, R³ and R⁴ are independently hydrogen, halo, —O(C₁-C₆ alkyl),—CON(R¹¹)₂, —SOR¹¹, —SO₂R¹¹, —SO₂N(R¹¹)₂, —N(R¹¹)₂, —NR¹¹COR¹¹,—CO₂R^(11 , —COR) ¹¹, —SR¹¹, —OH, —NO₂ or —CN; each R¹¹ is independentlyhydrogen or C₁-C₆ alkyl; R² is halogen; R⁵ and R⁶ are each independentlyhydrogen, or C₁-C₆ alkyl or together with the carbon atom to which theyare attached form a C₃-C₇ cycloalkyl group; R⁷ is methyl; R⁸ is phenyl,naphthalenvl, quinolinyl, quinoxalinyl, thiazolyl, or biphenyl, any ofwhich is optionally substituted with one or more substituents selectedfrom halo, C₁-C₆ alkyl, —O(C₁-C₆)alkyl, —CON(R¹¹)₂, —SOR¹¹, —SO₂R¹¹,—SO₂N(R¹¹)₂, —N(R¹¹)₂, —NR¹¹COR¹¹, —CO₂R¹¹, —COR¹¹, —SR¹¹, —OH, —NO₂ or—CN; wherein R¹¹ is as defined above; R⁹ is hydrogen, or C₁-C₆ alkyl;provided that: R⁸ is not unsubstituted phenyl or phenyl substituted with—COOH; said process comprising the hydrolysis of a compound of generalformula (II)

wherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, and R⁹ are as defined above andwherein R¹² is C₁-C₆ alkyl.
 2. A process for the preparation of acompound of general formula (II) as defined in claim 1, said processcomprising reacting a compound of general formula (III):

wherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷ and R¹² are as defined in claim 1;with a compound of general formula (IV):R⁹C(═O)R⁸   (IV) wherein R⁸ and R⁹ are as defined in claim 1; underacidic reductive alkylation conditions.
 3. A process for preparation ofa compound of general formula (I) according to claim 1

wherein R¹, R³ and R⁴ are hydrogen; R⁵ and R⁶ are hydrogen; R⁸ isquinolinyl; and R⁹ is hydrogen; said process comprising the hydrolysisof a compound of general formula (II)

wherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, and R⁹ are as defined above. 4.A process for the preparation of a compound of general formula (II) asdefined in claim 1, said process comprising reacting a compound ofgeneral formula (III):

wherein R¹, R³ and R⁴ are hydrogen; R⁵ and R⁶ are hydrogen; and R¹² isC₁-C₆ alkyl; with a compound of general formula (IV):R⁹C(═O)R⁸  (IV) wherein R⁸ is quinolinyl; and R⁹ is hydrogen; underacidic reductive alkylation conditions.
 5. The process of claim 1,wherein, independently or in combination: R¹ is halo or hydrogen; R³ ishalo or hydrogen; R⁴ is halo or hydrogen; R⁵ and R⁶ are eachindependently hydrogen or C₁-C₄ alkyl; and R⁹ is hydrogen or C₁-C₄alkyl.
 6. The process of claim 1, wherein at least one of R⁵ and R⁶ ishydrogen.
 7. The process of claim 1, wherein R⁸ is selected from thegroup consisting of phenyl substituted at the 4-position ornaphthalen-2-yl, quinolin-2-yl, quinoxalin-2-yl, or thiazol-2-yl, any ofwhich is optionally substituted with one or more of the substituentshalo, C₁-C₆ alkyl, —O(C₁-C₆ alkyl), —CON(R¹¹)₂, —SOR¹¹, —SO₂R¹¹,—SO₂N(R¹¹)₂, —N(R¹¹)₂, —NR¹¹COR¹¹, —CO₂R¹¹, —COR¹¹, —SR¹¹, —OH, —NO₂, or—CN; wherein each R¹¹ is independently hydrogen or C₁-C₆ alkyl.
 8. Theprocess of claim 1, wherein R⁸ is substituted with one or moresubstituents selected from the group consisting of halo, C₁-C₄ alkyl,C₁-C₄ haloalkyl, C₁-C₄ alkoxy, C₁-C₄ alkylsulfonyl, and hydroxy.
 9. Theprocess of claim 1, wherein R⁸ is substituted with one or moresubstituents selected from the group consisting of chloro, fluoro,methyl, ethyl, t-butyl, trifluoromethyl, methoxy, methanesulfonyl, andhydroxy.
 10. The process of claim 1, wherein the compound of generalformula (I) is selected from the group consisting of:{3-[1-(4-Chloro-phenyl)-ethyl]-5-fluoro-2-methyl-indol-1-yl}-aceticacid,{5-Fluoro-2-methyl-3-[1-(4-trifluoromethyl-phenyl)-ethyl]-indol-1-yl}-acetic acid,{3-[1-(4-tert-Butyl-phenyl)-ethyl]-5-fluoro-2-methyl-indo-1-yl}-aceticacid,{5-Fluoro-3-[1(4-methanesulfonyl-phenyl)-ethyl]-2-methyl-indol-1-yl}-aceticacid, [5-Fluoro-2-methyl-3-(1-naphthalen-2-yl-ethyl)-indol-1-yl]-aceticacid, (5-Fluoro-2-methyl-3-quinolin-2-ylmethyl-indol-1-yl)-acetic acid,(5-Fluoro-2-methyl-3-naphthalen-2-ylmethyl-indol-1-yl)-acetic acid,[5-Fluoro-3-(8-hydroxy-quinolin-2-ylmethyl)-2-methyl-indol-1-yl]-aceticacid, (5-Fluoro-2-methyl-3-quinoxalin-2-ylmethyl-indol-1-yl)-aceticacid, [5-Fluoro-3-(4-methoxy-benzyl)-2-methyl-indol-1-yl]-acetic acid,(5-Fluoro-2-methyl-3-thiazol-2-ylmethyl-indol-1-yl)-acetic acid ethylester, [3-(4-Chloro-benzyl)-5-fluoro-2-methyl-indol-1-yl]acetic acid,[5-Fluoro-2-methyl-3-(4-trifluoromethyl-benzyl)-indol-1-yl]-acetic acid,[5-Fluoro-2-methyl-3-(4-tert-butyl-benzyl)-indol-1-yl]-acetic acid,(3-Biphenyl-4-ylmethyl-5-fluoro-2-methyl-indol-1-yl)-acetic acid,[5-Fluoro-3-(4-methanesulfonyl-benzyl)-2-methyl-indol-1-yl]-acetic acid,[5-Fluoro-3-(6-fluoro-quinolin-2-ylmethyl)-2-methyl-indol-1-yl]-aceticacid, and (5-Chloro-2-methyl-3-quinolin-2-ylmethyl-indol-1-yl)-aceticacid.
 11. The process of claim 1, wherein the compound of generalformula (I) is(5-Fluoro-2-methyl-3-quinolin-2-ylmethyl-indol-1-yl)-acetic acid.